The WaterMaster™ DNA Purification Kit provides all of the reagents needed to purify microbial DNA from any water source after passage through a filter of your choice. This capability includes DNA from bacteria and eukaryotes. The DNA is suitable for restriction digests, PCR, subcloning, sequencing, or microarray analysis. The kit employs a simple, gentle procedure with no toxic organic reagents or mechanical cell disruption. The final purified DNA is in a 50-fold smaller volume than competing kits. Thus, there is no need to further concentrate the DNA.
As a test of the detection limit for bacteria in water samples, varying amounts of E. coli from 105 to 101 cells per 100 ml of water were filtered through 0.45 micron filters, DNA was extracted with the WaterMaster Kit, and detected by PCR (Fig. 1). It is evident in this figure that the DNA from as few as 10 bacteria per 100 ml of water can be extracted by the WaterMaster Kit and detected by PCR using primers specific for a eubacterial rRNA gene.1
Figure 1. Detection of E. coli in water samples by PCR. Varying amounts of E.coli K12 were diluted in 100-ml distilled water samples, trapped on 0.45-micron filters and DNA was extracted using the WaterMaster™ Kit. Aliquots of the extracted DNA were used as templates for PCR, using primers for a eubacterial rRNA gene and the FailSafe™ PCR System. The PCR products were analyzed by agarose gel electrophoresis. Lane M, 100-bp ladder; PCR products from DNA purified from the indicated amounts of E. coli cells: lane 1, 105; lane 2, 103; lane 3, 102; lane 4, 101; lane 5, negative control. The amplicon is 350 bp.
Figure 2. High-molecular-weight DNA was purified from microbes in a water sample from a spring-fed pond using the WaterMaster™ DNA Purification Kit. The 1% agarose gel was stained with SYBR® Gold. Lane M, Kilobase DNA ladder; lane 1, a 5-ng aliquot of DNA purified with the WaterMaster Kit.